This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The goal of the project is to determine the mechanisms of peptide loading onto human MHC class II molecules. MHC class II molecules bind peptides very tightly, with half-lives of several days to weeks. Empty MHC class II molecules have a tendency to aggregate and they assemble in the ER with invariant chain, which occupies the binding groove like a peptide. In the endosomal peptide loading compartment, invariant chain is degraded by a set of proteases, leaving the CLIP peptide of invariant chain in the groove. Removal of the CLIP peptide requires the action of HLA-DM and makes the binding groove vacant for binding of microbial peptides. We have defined the molecular requirements for the interaction of MHC class II molecules with HLA-DM. We found that HLA-DM binding required release of the N-terminal part of the peptide. Based on this insight, we are now attempting to characterize this interaction at a structural level.